The overall goal of this revised project is to determine the effect of cannabinoids and the role CB2 plays in the regulation of B cell function in order to better understand the immunomodulatory potential of cannabinoid-based drugs and the role of the endocannabinoid system in immune homeostasis. The world-wide use of marijuana and other cannabinoids in the treatment of chronic diseases is on the increase making it extremely important to understand the full immunological effects of these drugs. Cannabinoids and related compounds have been proposed for use in treating rheumatoid arthritis and irritable bowel syndrome and have actually been used in clinical trials for brain injury, multiple sclerosis and HIV-induced wasting disease. It is now clear from previous studies that CB2 receptors are expressed on B cells and that cannabinoids regulate B cell function; therefore, to increase our understanding of the immunological consequences of cannabinoid use we propose the following specific aims. Aim 1. To determine in mouse immunization models the effect of non-selective cannabinoid agonists on Th2-biasing antibody production in response to standardized antigens such as NP-OVA and KLH as well as to the Th1-dependent infectious agent, Listeria monocytogenes. Antibody production and Th2-dependent isotype switching will be determined by ELISA of serum samples and by examining T helper biasing cytokine production in spleen and lymph node cells from injected mice. We hypothesize that cannabinoids are immune adjuvants selective for antibody production by Th2 biasing mechanisms. Aim 2. To Determine if cannabinoids directly stimulate in vitro B cell activation and maturation. Mouse splenic B cells will be purified and cultured with cannabinoids and B cell activators such as IL-4 and anti-CD-40. B cell proliferation will be determined by the CyQuant assay. B cell maturation will be assessed by flow cytometry analysis of the IgM to IgE class switch, and B cell activation will be assessed by the expression of cell surface activation markers. We hypothesize that cannabinoids can directly modulate B cell activation and maturation independent of T cell help. Aim 3. To determine the role of CB2 in cannabinoid modulation of antibody production and other B cell functions; also determine if B cell activation increases the expression of CB2 and if ligation of B cell receptors such as CD40, IL-4R, Toll-like, and CB2 produces overlapping transcription factor induction. We will use agonists and antagonists selective for CB2 as well as knockout mice to determine the role of this receptor in the drug effects on B cell function. Also, CB2 mRNA expression will be analyzed in stimulated B cell cultures by RT coupled with real time PCR; expression of overlapping transcription factors will be studied by commercially available gene array kits.